Heterologous Expression of Fungal Cellobiohydrolase 2 Genes in Yeast

Abstract

The present invention provides for heterologous expression of polypeptides encoded by wild-type and codon-optimized cbh2 genes from the organisms Cochliobolus heterostrophus, Gibberella zeae, Irpex lacteus, Volvariella volvacea, and Piromyces sp. in host cells, such as the yeast Saccharomyces cerevisiae. The expression in such host cells of the corresponding genes, and variants and combinations thereof, result in improved specific activity of the expressed cellobiohydrolases. Thus, such genes and expression systems are useful for efficient and cost-effective consolidated bioprocessing systems.

Description

BACKGROUND OF THE INVENTION[0001]Lignocellulosic biomass is widely recognized as a promising source of raw material for production of renewable fuels and chemicals. The primary obstacle impeding the more widespread production of energy from biomass feedstocks is the general absence of low-cost technology for overcoming the recalcitrance of these materials to conversion into useful fuels. Lignocellulosic biomass contains carbohydrate fractions (e.g., cellulose and hemicellulose) that can be converted into ethanol. In order to convert these fractions, the cellulose and hemicellulose must ultimately be converted or hydrolyzed into monosaccharides; it is the hydrolysis that has historically proven to be problematic.[0002]Biologically mediated processes are promising for energy conversion, in particular for the conversion of lignocellulosic biomass into fuels. Biomass processing schemes involving enzymatic or microbial hydrolysis commonly involve four biologically mediated transformation...

AI Technical Summary

Benefits of technology

The present patent provides a method for efficiently breaking down cellulosic materials such as plant matter into sugar for the production of ethanol. This is achieved by introducing specific genes from different microorganisms into a host cell. The use of these genes helps to improve cellulose hydrolysis and enhance ethanol production in the host cells.

Problems solved by technology

The primary obstacle impeding the more widespread production of energy from biomass feedstocks is the general absence of low-cost technology for overcoming the recalcitrance of these materials to conversion into useful fuels.

In order to convert these fractions, the cellulose and hemicellulose must ultimately be converted or hydrolyzed into monosaccharides; it is the hydrolysis that has historically proven to be problematic.

First, Fujita et al. were unable to achieve fermentation of amorphous cellulose using yeast expressing only recombinant Bgl1 and EgII.

A second limitation of the Fujita et al. approach was that cells had to be pre-grown to high cell density on standard carbon sources before the cells were useful for ethanol production using amorphous cellulose (e.g., Fujita et al. uses high biomass loadings of ˜15 g / L to accomplish ethanol production).

However, current levels of expression and specific activity of cellulases heterologously expressed in yeast are still not sufficient to enable growth and ethanol production by yeast on cellulosic substrates without externally added enzymes.

Currently, there is no reliable way to predict which cellulases will be efficiently expressed in heterologous organisms.

Furthermore, even cellulases which are expressed at high levels may not be particularly active in a heterologous organism.

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